Responses of skeletal muscle size and anabolism are reproducible with multiple periods of unloading/reloading.

Mechanical unloading has long been understood to contribute to rapid and substantial adaptations within skeletal muscle, most notably, muscle atrophy. Studies have often demonstrated that many of the alterations resulting from disuse are reversed with a reintroduction of load and have supported the concept of muscle plasticity. We hypothesized that adaptations during disuse and recovery were a repeatable/reproducible phenomenon, which we tested with repeated changes in mechanical load. Rats were assigned to one of the following five groups: animals undergoing one or two bouts of hindlimb unloading (28 days), with or without recovery (56 day), or control. Following the completion of their final time point, posterior crural muscles were studied. Muscle sizes were lower following 28 days of disuse but fully recovered with a 56-day reloading period, regardless of the number of disuse/recovery cycles. Mixed protein fractional synthesis rates consistently reflected mass and loading conditions (supported by anabolic signaling), whereas the myofibrillar protein synthesis response varied among muscles. Amino acid concentrations were assessed in the gastrocnemius free pool and did not correlate with muscle atrophy associated with mechanical unloading. Muscle collagen concentrations were higher following the second unloading period and remained elevated following 56 days of recovery. Anabolic responses to alterations in load are preserved throughout multiple perturbations, but repeated periods of unloading may cause additive strain to muscle structure (collagen). This study suggests that whereas mass and anabolism are reproducibly reflective of the loading environment, repeated exposure to unloading and/or reloading may impact the overall structural integrity of muscle. NEW & NOTEWORTHY Repeatability should be considered a component of skeletal muscle plasticity during atrophy and recovery. Muscle anabolism is equally affected during a first or second disuse bout and returns equally with adequate recovery. Elevated muscle collagen concentrations observed after the second unloading period suggest altered structural integrity with repeated disuse.

Larval development of the Mexican Snook, Centropomus poeyi (Teleostei: Centropomidae)

We document for the first time the early ontogeny of Centropomus poeyi based on captive raised material representing 0-19 days posthatch (dph). The achievement of early developmental landmarks (i.e., yolk-sac depletion, flexion, development of fins) and changes in pigmentation are described (1.4 mm NL-10.6 mm SL; 0-19 dph) and documented for a subset of individuals using high quality photographs. The ontogeny of the viscerocranium is also described (2.4 mm NL-10.6 mm SL; 6-19 dph). Development in C. poeyi occurs over a short period with attainment of the juvenile stage (i.e., full complement of fin rays present in each fin) occurring by 6.9 mm SL. The ontogeny of external pigmentation in C. poeyi is marked by two trends throughout growth: (1) a decrease in pigmentation dorsally; and (2) an increase in pigmentation ventrally along the midline. Development of the viscerocranium begins with the appearance of the maxilla and dentary in individuals of 2.4 mm NL, coinciding with the depletion of the yolk-sac. By 10.6 mm SL all bones of the viscerocranium are present and teeth are present on all teeth-bearing bones of the adult. Aspects of early development in C. poeyi are compared with the congeners C. undecimalis and C. parallelus.(AU)

Se documentó por primera vez la ontogenia temprana de Centropomus poeyi basada en material obtenido de cautiverio a partir de embriones (0 días después de la eclosión, dde) hasta los 19 dde. Se registró la aparición de puntos de referencia en el desarrollo temprano (p.ej. consumo de saco vitelino, flexión, desarrollo de aletas) y los cambios de pigmentación han sido descritos (1.4 mm NL-10.6 mm SL; 0-19 dde), siendo documentado para un subgrupo de individuos utilizando fotografías de alta calidad. El desarrollo del viscerocráneo también fue descrita (2.4 mm NL-10.6 mm SL; 6-19 dde). El desarrollo en C. poeyi sucede en un periodo corto de tiempo hasta alcanzar una etapa juvenil (p.ej., aparición completa de los radios presentes en cada aleta), lo que se observa a los 6.9 mm SL. La pigmentación externa en C. poeyi esta marcada por dos tendencias en el crecimiento: (1) un descenso en la pigmentación dorsal; y (2) un incremento en la pigmentación ventral a lo largo de la línea media. El desarrollo del viscerocráneo comienza con el aparecimiento del maxilar y el dentario en individuos de 2.4 mm NL, lo cual coincide con el consumo del saco vitelino. A los 10.6 mm SL todos los huesos del viscerocráneo están presentes como se observa en los dientes en los adultos. Algunos aspectos del desarrollo temprano en C. poeyi son comparados con los congéneros C. undecimalis y C. parallelus.(AU)

Expressed IgH µ and τ transcripts share diversity segment in ranched Thunnus orientalis.

It is now appreciated that in addition to the immunoglobulin (Ig)M and D isotypes fish also make the mucosal IgT. In this study we sequenced the full length of Ig τ as well as µ in the commercially important Thunnus orientalis (Pacific bluefin tuna), the first molecular analysis of these two Ig isotypes in a member of the order Perciformes. Tuna IgM and IgT are each composed of four constant (CH) domains. We cloned and sequenced 48 different variable (VH) domain gene rearrangements of tuna immunoglobulins and grouped the VH gene sequences to four VH gene segment families based on 70% nucleotide identity. Three VH gene families were used by both IgM and IgT but one group was only found to be used by IgM. Most interestingly, both µ and τ clones appear to use the same diversity (DH) segment, unlike what has been described in other species, although they have dedicated IgT and IgM joining (JH) gene segments. We complemented this repertoire study with phylogenetic and tissue expression analysis. In addition to supporting the development of humoral vaccines in this important aquaculture species, these data suggest that the DH-JH recombination rather than the VH-DH recombination may be instructive for IgT versus IgM/D bearing lymphocyte lineages in some fish.

The effect of aflatoxin-B1 on red drum (Sciaenops ocellatus) and assessment of dietary supplementation of NovaSil for the prevention of aflatoxicosis.

Aflatoxin B1 (AFB1) is a potent carcinogen that causes growth stunting, immunosuppression and liver cancer in multiple species. The recent trend of replacing fishmeal with plant-based proteins in fish feed has amplified the AFB1 exposure risk in farm-raised fish. NovaSil (NS), a calcium montmorillonite clay, has previously been shown to reduce AFB1 bioavailability safely and efficaciously in several mammalian species. This study was designed to: (1) evaluate AFB1 impact on cultured red drum, Sciaenops ocellatus, over the course of seven weeks; and (2) assess NS supplementation as a strategy to prevent aflatoxicosis. Fish were fed diets containing 0, 0.1, 0.25, 0.5, 1, 2, 3, or 5 ppm AFB1. Two additional treatment groups were fed either 5 ppm AFB1 + 1% NS or 5 ppm AFB1 + 2% NS. Aflatoxin B1 negatively impacted red drum weight gain, survival, feed efficiency, serum lysozyme concentration, hepatosomatic index (HSI), whole-body lipid levels, liver histopathological scoring, as well as trypsin inhibition. NovaSil inclusion in AFB1-contaminated diets improved weight gain, feed efficiency, serum lysozyme concentration, muscle somatic index, and intraperitoneal fat ratios compared to AFB1-treated fish. Although not significant, NS reduced AFB1-induced histopathological changes in the liver and decreased Proliferating Cell Nuclear Antigen (PCNA) staining. Importantly, NS supplementation improved overall health of AFB1-exposed red drum.

Effects of dietary arginine on endocrine growth factors of channel catfish, Ictalurus punctatus.

Thyroid (TH) and growth (GH) hormones, and insulin-like growth factor I (IGF-I) are anabolic regulators in fish and responsive to nutrient intake. A study was conducted to determine if previously reported growth effects of dietary arginine (ARG) in channel catfish were related to the activation of endocrine axes. In a first experiment, catfish were fed incremental levels of ARG (0.5 - 4% of diet) for 6 weeks and sampled at 2-week intervals. In a second experiment, fasted (48h) fish were fed a single ration of ARG (0.5 or 4% of diet) and sampled at various intervals (0 to 72h postprandial, PP). Experiment 1 did not reveal any influence of ARG on circulating TH, GH, or IGF-I despite the significantly increased growth of fish fed ARG-enriched diets. In experiment 2, feeding the 4% ARG diet significantly increased the amplitude of pulsatile plasma GH levels and also significantly increased IGF-I mRNA in liver and muscle, (at 2h PP) and plasma IGF-I levels (at 6h PP). Although relatively infrequent sampling failed to reveal alterations in TH or GH levels in response to ARG-induced growth activation, PP high frequency sampling unveiled high amplitude pulsatile GH secretions and may be important in activating IGF production in target tissues. Additionally, expressed and secreted IGF-I exhibited discernible patterns which closely correlate with ARG-induced growth effects in catfish.

Mechanisms of intrinsic resistance to antimicrobial peptides of Edwardsiella ictaluri and its influence on fish gut inflammation and virulence.

The genus Edwardsiella comprises a genetically distinct taxon related to other members of the family Enterobacteriaceae. It consists of bacteria differing strongly in their biochemical and physiological features, natural habitats, and pathogenic properties. Intrinsic resistance to cationic antimicrobial peptides (CAMPs) is a specific property of the genus Edwardsiella. In particular, Edwardsiella ictaluri, an important pathogen of the catfish (Ictalurus punctatus) aquaculture and the causative agent of a fatal systemic infection, is highly resistant to CAMPs. E. ictaluri mechanisms of resistance to CAMPs are unknown. We hypothesized that E. ictaluri lipopolysaccharide (LPS) plays a role in both virulence and resistance to CAMPs. The putative genes related to LPS oligo-polysaccharide (O-PS) synthesis were in-frame deleted. Individual deletions of wibT, gne and ugd eliminated synthesis of the O-PS, causing auto-agglutination, rough colonies, biofilm-like formation and motility defects. Deletion of ugd, the gene that encodes the UDP-glucose dehydrogenase enzyme responsible for synthesis of UDP-glucuronic acid, causes sensitivity to CAMPs, indicating that UDP-glucuronic acid and its derivatives are related to CAMP intrinsic resistance. E. ictaluri OP-S mutants showed different levels of attenuation, colonization of lymphoid tissues and immune protection in zebrafish (Danio rerio) and catfish. Orally inoculated catfish with O-PS mutant strains presented different degrees of gut inflammation and colonization of lymphoid tissues. Here we conclude that intrinsic resistance to CAMPs is mediated by Ugd enzyme, which has a pleiotropic effect in E. ictaluri influencing LPS synthesis, motility, agglutination, fish gut inflammation and virulence.

The effects of prebiotics on the digestive enzymes and gut histomorphology of red drum (Sciaenops ocellatus) and hybrid striped bass (Morone chrysops × M. saxatilis).

The effects of four prebiotics (fructo-oligosaccharide, Bio-MOS, transgalacto-oligosaccharide and GroBiotic-A) on digestive enzymes and intestinal morphology were studied in juvenile hybrid striped bass (Morone chrysops × M. saxatilis) and red drum (Sciaenops ocellatus) using two separate 8-week feeding trials. Red drum were fed experimental diets with the four prebiotics each individually supplemented at 1% and hybrid striped bass were fed diets supplemented with GroBiotic-A at 1 and 2%. Both trials were conducted with each diet fed to apparent satiation twice per d to three replicate groups of fifteen juvenile fish. For histomorphological analysis, gastrointestinal tract (GIT) samples from three randomly selected fish per tank were taken at 4 and 8 weeks for hybrid striped bass and at 8 weeks for red drum. For both trials, GIT samples from two randomly selected fish per tank were taken at 4 and 8 weeks and analysed for pepsin, trypsin, chymotrypsin, aminopeptidase, α-amylase, lipase, and both acid and alkaline phosphatase activities. The results of the histological evaluation indicated that the inclusion of prebiotics was adequate to elicit structural changes in the GIT of both species. On the other hand, no significant changes in the enzyme activities were detected at week 8 in both species. However, there was a transient effect of Bio-MOS supplementation on the activities of aminopeptidase, α-amylase and alkaline phosphatase at week 4 in red drum only. Thus, previously observed improvements in nutrient digestibility by these fish in response to prebiotic supplementation appear to be mostly related to changes in GIT structure as opposed to the enhancement of digestive enzyme activity.

Synergies between vaccination and dietary arginine and glutamine supplementation improve the immune response of channel catfish against Edwardsiella ictaluri.

Channel catfish was used to investigate the enhancement of vaccine efficacy following dietary supplementation with arginine (ARG, 4% of diet), glutamine (GLN, 2% of diet), or a combination of both. After vaccination against Edwardsiella ictaluri, humoral and cellular immune responses, along with lymphoid organ responses were evaluated. E. ictaluri-specific antibody titers in plasma were higher (P < 0.05) in fish fed the supplemented diets compared to those fed the basal diet as early as 7 d post-vaccination (dpv). B-cell proportion in head-kidney was higher (P < 0.05) at 14 dpv in vaccinated fish fed the GLN diet. The responsiveness of spleen and head-kidney lymphocytes against E. ictaluri was enhanced (P < 0.05) by dietary supplementation of ARG or GLN at 14 dpv. Additionally, at 7 dpv, vaccinated fish fed the GLN diet had higher (P < 0.05) head kidney weights relative to the other dietary treatments, and vaccinated fish fed ARG-supplemented diets had higher (P < 0.05) protein content in this tissue. Results from this study suggest that dietary supplementation of ARG and GLN may improve specific cellular and humoral mechanisms, enhancing the acquired immunity in vaccinated channel catfish.

Arginine and glutamine supplementation to culture media improves the performance of various channel catfish immune cells.

Specific components of both the innate and adaptive immune systems of channel catfish were evaluated after supplementation of culture media with arginine (ARG) and/or glutamine (GLN). Primary cell cultures of head-kidney macrophages (MØ) were used for phagocytic and bactericidal assays against Edwardsiella ictaluri. Additionally, proliferation assays were conducted with naïve peripheral blood lymphocytes (PBL) exposed to non-specific mitogens. To indirectly assess amino acid utilization of both MØ and PBL, amino acid levels, with emphasis on ARG and GLN, were evaluated in the basal medium before and after activation or mitogenic exposure. After bactericidal and proliferation assays, the sum of the media free amino acid pool significantly (P < 0.05) decreased 23% and 45%, respectively. Glutamine levels in medium decreased by 38% and ARG by 18% during the bactericidal assay. Also, decreases of 52 and 46% from initial values were found after the proliferation assay for GLN and ARG, respectively. Macrophage phagocytosis and killing ability was significantly (P < 0.05) enhanced by ARG supplementation to culture media regardless of GLN supplementation. Proliferation of naïve T- and B-lymphocytes upon mitogenic exposure was significantly (P < 0.05) enhanced by supplementing ARG and GLN to the media, but limited synergistic effects were observed. These results suggest that in vitro, ARG and GLN are important substrates and immunomodulators of both innate and adaptive responses in fish leukocytes, and further highlights the potential use of ARG and GLN as immunonutrients in aquafeeds.

A novel approach for assessing protein synthesis in channel catfish, Ictalurus punctatus.

A comprehensive understanding of animal growth requires adequate knowledge of protein synthesis (PS), which in fish, has traditionally been determined by the flooding dose method. However, this procedure is limited to short-term assessments and may not accurately describe fish growth over extended periods of time. Since deuterium oxide ((2)H(2)O) has been used to non-invasively quantify PS in mammals over short- and long-term periods, we aimed at determining if (2)H(2)O could also be used to measure PS in channel catfish. Fish were stocked in a 40-L aquarium with approximately 4% (2)H(2)O and sampled at 4, 8 and 24h (n=6 at each time period) to determine (2)H-labeling of body water (plasma), as well as protein-free and protein-bound (2)H-labeled alanine. The labeling of body water reflected that of aquarium water and the labeling of protein-free alanine remained constant over 24h and was approximately 3.8 times greater than that of body water. By measuring (2)H-labeled alanine incorporation after 24h of (2)H(2)O exposure we were able to calculate a rate of PS: 0.04+/-0.01% h(-1). These results demonstrate that PS in fish can be effectively measured using (2)H(2)O and, because this method yields integrative measures of PS, is relatively inexpensive and accounts for perturbations such as feeding, it is a novel and practical assessment option.